We propose to study the mechanisms whereby HIV cDNA becomes joined to host cell DNA, a process that serves as a model of DNA breaking and joining reactions. It is possible to infect cells with HIV, then isolate integration competent complexes containing the viral cDNA associated with viral and cellular proteins (preintegration complexes; "PICs"). We will develop new high resolution footprinting methods to study PICs employing a novel Tn7 based transposase system. This method is needed because PICs cannot be isolated in large quantities, placing additional demands on footprinting technology. We will purify PICs, and characterize their protein composition and architecture. Already we have carried out high resolution and high sensitivity footprinting reactions on a well studied model system, and are beginning to apply these methods to integration competent replication intermediates. We will also develop new DNA substrates to probe the DNA architecture required for integration, and to serve as co-crystallization reagents in structural studies. These studies will help lead to the development of novel antiretroviral agents which will inhibit integration.